Metabolic-Associated Fatty Liver Disease (MAFLD), Diabetes, and Cardiovascular Disease: Associations with Fructose Metabolism and Gut Microbiota

Non-alcoholic fatty liver disease (NAFLD) is an increasingly common condition associated with type 2 diabetes (T2DM) and cardiovascular disease (CVD). Since systemic metabolic dysfunction underlies NAFLD, the current nomenclature has been revised, and the term metabolic-associated fatty liver disease (MAFLD) has been proposed. The new definition emphasizes the bidirectional relationships and increases awareness in looking for fatty liver disease among patients with T2DM and CVD or its risk factors, as well as looking for these diseases among patients with NAFLD. The most recommended treatment method of NAFLD is lifestyle changes, including dietary fructose limitation, although other treatment methods of NAFLD have recently emerged and are being studied. Given the focus on the liver–gut axis targeting, bacteria may also be a future aim of NAFLD treatment given the microbiome signatures discriminating healthy individuals from those with NAFLD. In this review article, we will provide an overview of the associations of fructose consumption, gut microbiota, diabetes, and CVD in patients with NAFLD.

The definition of NAFLD combines the presence of steatosis in more than 5% of hepatocytes and metabolic risk factors, especially obesity and T2DM, and exclusion of excessive alcohol consumption defined as ≥30 g per day for men and ≥20 g per day for women, or other chronic liver diseases [9]. It should be noted that the liver is a primary organ for lipid and glucose homeostasis and is the focus of cardiometabolic disease. In 2020, fatty liver was redefined from negative (absence of excessive alcohol consumption and other known causes of liver disease) to a more positively stated metabolic associated fatty liver disease (MAFLD) [10]. The latter definition of MAFLD is based on the presence of hepatic steatosis and at least one other condition such as overweight/obesity, T2DM, or metabolic abnormalities with no additional exclusion criteria [10,11]. Metabolic abnormalities included in the definition cover at least two features from the following: increased waist circumference, arterial hypertension, hypertriglyceridemia, low high-density cholesterol (HDL-C), prediabetes, insulin resistance, and subclinical inflammation [10] (Figure 2). This new definition underlines the importance of cardiometabolic risk factors contributing to liver disease even among patients with other liver diseases and who drink alcohol [10]. Although it should be noticed that although the new definition has been proposed by a panel of international experts from 22 countries, this new nomenclature is not yet accepted by the American Association for the Study of Liver Disease and the European Association for the Study of Liver Disease.
Diabetes mellitus is a heterogeneous group of disorders that results in an increase in blood glucose concentration, yet the pathophysiological processes underlying type 1 and type 2 of the disease differ. Insulin resistance, which is tightly linked to T2DM, is not always present in type 1 of the disease, thereby explaining the different prevalence of NAFLD in those two subpopulations of patients [11]. The latter definition of MAFLD is based on the presence of hepatic steatosis and at least one other condition such as overweight/obesity, T2DM, or metabolic abnormalities with no additional exclusion criteria [10,11]. Metabolic abnormalities included in the definition cover at least two features from the following: increased waist circumference, arterial hypertension, hypertriglyceridemia, low high-density cholesterol (HDL-C), prediabetes, insulin resistance, and subclinical inflammation [10] (Figure 2). This new definition underlines the importance of cardiometabolic risk factors contributing to liver disease even among patients with other liver diseases and who drink alcohol [10]. Although it should be noticed that although the new definition has been proposed by a panel of international experts from 22 countries, this new nomenclature is not yet accepted by the American Association for the Study of Liver Disease and the European Association for the Study of Liver Disease.
Diabetes mellitus is a heterogeneous group of disorders that results in an increase in blood glucose concentration, yet the pathophysiological processes underlying type 1 and type 2 of the disease differ. Insulin resistance, which is tightly linked to T2DM, is not always present in type 1 of the disease, thereby explaining the different prevalence of NAFLD in those two subpopulations of patients [11].

NAFLD and Fructose
Fructose's use in foods was limited until the late 1960s due to its high price [25]. Since then, it has become freely available and has been shown to exert a positive effect in the treatment of diabetes since fructose does not require insulin to be metabolized and has no effect on fasting blood glucose levels and urinary glucose excretion [26,27]. Nowadays, the way scientists look at fructose has changed, and it is now known as a risk factor in the development of obesity and several metabolic disturbances, NAFLD, among others [19].
Fructose is a major dietary monosaccharide that occurs naturally in ripe fruits, honey, and in small amounts in some vegetables such as carrot, onion, paprika, and sweet potato.
It also comes in industrially manufactured foods because it is a main ingredient in the most widely used sweeteners like disaccharide sucrose (table sugar, composed of one glucose molecule and one fructose) and high fructose glucose syrup (mixture of fructose with sucrose or glucose) [18,27].
Fructose, in contrast to glucose, is almost totally cleared from circulation by the liver with the use of glucose transporter type-5 (GLUT 5). A large amount of acetyl-CoA is produced following fructose uptake because fructose clearance omits glycolysis, which is the rate-limiting step in acetyl-CoA production [28]. Some acetyl-CoA is used for ATP production, but the excess amount is used for de novo lipogenesis, which is one of the mechanisms proposed for how consuming fructose leads to NAFLD [28,29]. Based on the results of studies on animals and humans with fructose overfeeding, high fructose consumption (25% of energy requirement) may increase visceral adiposity, postprandial hypertriglyceridemia, and insulin resistance by acting on de novo lipogenesis [30].
However, it is not only lipogenesis; other hepatotoxic effects are exerted by fructose, namely inducing an increase in oxidative stress [31]. Fructose is able to directly generate reactive oxygen species (ROS) and lead to hepatocellular damage through protein fructosylation [32]. Because of the increased consumption of processed foods, fructose consumption has increased dramatically, by 30% over the last 40 years and by 500% over the last century [33]. Also, there was more than a 40% increase in the intake of sugar-sweetened beverages (SSB) from 1990 to 2016 [34]. Because free sugars consumption has a proven association with metabolic diseases and cancer, it has since been recommended by the World Health Organization (WHO) to decrease its consumption to less than 10% of total daily energy intake [35].
Numerous animal and human studies have revealed the association of a close relationship between the consumption of fructose and the development of NAFLD (Table 1). There was no effect of fructose in isocaloric trials. Increased consumption of fructose in hypercaloric trials increased both IHCL and ALT. Abbreviations: ALT-alanine aminotransferase, AST-aspartate aminotransferase, CI-credible interval, CRP-Creactive protein, DNL-de novo lipogenesis, HDL-c-high-density lipoprotein cholesterol, LDL-c-low-density lipoprotein cholesterol, IHCL-intrahepatocellular lipids, NAFLD-non-alcoholic fatty liver disease, MD-mean difference, OR-odds ratio, RCT-randomized controlled trial, RR-risk ratio, SMD-standardized mean difference, SSB-sugar-sweetened beverages, SSSB-sucrose-sweetened soft drinks.
In a systematic review and meta-analysis of observational studies involving 4639 individuals, SSB consumption led to a 53% increased risk of developing NAFLD in comparison to participants who did not ingest SSB [46]. Another analysis of 12 studies with 35,705 individuals showed that higher consumption of SBB was associated with a 40% increase in the incidence of NAFLD [47]. Other studies also reported increased risk of NAFLD associated with consumption of SBB [36][37][38][39][40]43], yet some of the meta-analyses indicate the increase of intrahepatocellular lipids only under conditions of hypercaloric diet [41,42]. An interesting observation is that the association between SBB intake and liver fat may be independent of BMI [45]. One small RCT concluded that there was no significant change in hepatic fat or body weight in the groups consuming fructose or glucose beverages only; however, reduction of fructose led to improvement of several factors related to CVD [44]. What is also interesting to note is that fructose consumption influences the composition and function of gut microbiota in a way that promotes the development and progression of NAFLD [48].

NAFLD and Gut Microbiota
The pioneering studies using germ-free mice and gut microbiota transfer related to the association of gut microbiome with metabolic diseases revealed a contribution of gut microbiota to weight gain and metabolic alterations [49]. During the last decade, there has been a growing body of evidence demonstrating the contribution of the gut microbiome to the pathogenesis of NAFLD [50][51][52]. In general, dysfunction of the gut-liver axis caused by bacterial proliferation in the intestine, alteration of the intestinal permeability, and intestinal dysbiosis have a large influence on the development and progression of NAFLD [11]. Initial studies demonstrated that genetically modified mice (modification in the inflammasome pathway) were prone to develop NASH when co-housed with wild-type mice, leading to the development of liver steatosis and inflammation in wild-type mice as a consequence of microbiota sharing through coprophagia [53].
Fecal microbiota transfer from patients with NASH to germ-free mice causes hepatic steatosis and inflammation in these animals [54]. On the other hand, the outcomes of animal studies cannot be directly extrapolated to humans (e.g., mice do not develop the whole range of steatosis stages seen in humans, and mice microbiota itself differs from humans) [55,56].
Human studies have been based on comparisons of gut microbiota between patients with NAFLD, NASH, and cirrhosis and individuals with a healthy liver performed to demonstrate gut microbiota signatures in these pathological conditions [57]. Gut microbiota, and specifically, the gut-liver axis' role in the pathogenesis of NAFLD, has been explored in up-to-date studies, but this relationship is still poorly defined. Nevertheless, gut microbiome signatures in NAFLD, NAFLD fibrosis, and cirrhosis could become non-invasive diagnostic biomarkers for liver disease diagnosis [21].
The gut-liver axis is an association between gut microbiota and the liver. The interaction is conducted through the portal vein, which transports products from the gut to the liver and, in return, bile and antibodies from the liver to the intestine [58]. An important determining health factor seems to be a mucosal barrier comprised of intestinal epithelial cells. Its permeability and mucus composition are derived from gut microbiota and the presence of immune cells [58,59]. Increased permeability of the intestinal mucosal barrier and unfavorable changes in gut microbiota compositions are possible factors in the development and progression of NAFLD [58,59]. Dysfunction of gut microbiota results in the production of PAMPs (pathogen-associated molecular patterns), and increased permeability of the mucosal barrier leads to increased inflammation in the liver and the development and progression of liver disease [58][59][60]. Studies have shown a lower diversity of microbiota in patients with NAFLD compared to healthy controls [61,62].
Abnormalities of gut microbiota composition in stools of patients with NAFLD have been highlighted in a meta-analysis showing increased abundance of Escherichia, Prevotella, and Streptococcus and decreased abundance of Coprococcus, Feacalibacterium, and Ruminococcus [63]. When patients with liver fibrosis and patients with severe steatosis without fibrosis were compared, fecal Clostridium was significantly decreased in patients with liver fibrosis and was negatively associated with liver stiffness measurement (LSM) and myosteatosis [64]. Patients with liver fibrosis had increased fecal abundance of Escherichia and Shigella compared to patients with severe steatosis without fibrosis [64]. In another study in patients with NASH, there were increased levels of Escherichia and Shigella comparing patients with liver biopsy results as F0 (absence of fibrosis) or F1 (perisinusoidal or periportal fibrosis) [62].
NAFLD also results in increased fecal ester volatile organic compounds (VOC) and the presence of higher concentrations of fecal propionate and isobutyric acid and serum 2-hydroxybutyrate and L-lactic acid [65,66]. VOC is the result of gut microbiota substrate fermentation and is considered a potential marker of intestinal dysbiosis [67]. The results of the mentioned studies are summarized in Table 2. Abbreviations: CI-credible interval, NAFLD-non-alcoholic fatty liver disease, NASH-non-alcoholic steatohepatitis, SMD-standardized mean difference, VOC-volatile organic compounds.
According to the definitions formulated by the WHO and Food and Agriculture Organization of the United Nations (FAO), probiotics are live strains of strictly selected microorganisms which, when administered in adequate amounts, confer a health benefit on the host [73]. Prebiotics are described as a nonviable food component that confers a health benefit on the host associated with modulation of microbiota [74]. Synbiotics are the combination of synergistically acting probiotics and prebiotics; their role is the improvement of the survival of probiotic microorganisms in the gastrointestinal tract [75].
In this review article, we focus on the meta-analysis of RCT (randomized controlled trials), showing that probiotic/synbiotic therapy results in improving liver enzymes' activity and/or reduced steatosis/fibrosis in patients with NAFLD [68][69][70][71]. Moreover, treatment with probiotics decreases levels of CRP (C-reactive protein) and TNF-α (tumor necrosis factor α), suggesting the reduction of inflammation and playing an important role in NAFLD pathogenesis [68,70,72,76]. The studies related to synbiotics in patients with NAFLD are limited. In a recent RCT, there was a reduction in steatosis and improved liver enzyme changes observed in patients treated with bifidobacterium animalis and insulin [77]. In another study, the use of synbiotics positively influenced inflammatory markers in NAFLD [78].
The role of FMT in patients with NAFLD has limited RCT evidence. FMT was first proven to be a good treatment method of antibiotic-resistant clostridium difficile infection in 2013 [79] and soon after was tested in other diseases, including metabolic diseases [80]. In an RCT including 21 patients undergoing allogenic or autologous FMT, the procedure was not associated with an improvement in insulin resistance nor hepatic proton density fat fraction but had the potential to lower small intestinal permeability [81]. In the study by Witjes et al., allogenic FMT using lean vegan donors modified gut microbiota composition with favorable changes in plasma metabolites and steatohepatitis [82]. The aforementioned studies are summarized in Table 3. Allogenic FMT was associated with modified gut microbiota composition (increased abundance of ruminococcus, eubacterium hallii, faecalibacterium, and prevotella copri), decreased levels of GGT, a trend toward improvement in the necro-inflammation score (consisting of both lobular inflammation and hepatocellular ballooning). There was no significant difference in fibrosis score after allogenic FMT.

NAFLD and T2DM
According to epidemiological data, the overall prevalence of NAFLD among patients with T2DM is 55.5%, and NASH is 37.3%; also, 17% of T2DM patients who underwent liver biopsy have advanced fibrosis [14]. Current guidelines emphasize the role of screening for diabetes in patients diagnosed with NAFLD and vice versa [9,83]. NAFLD itself increases the risk of T2DM incidence [84][85][86], and the risk of the new-onset T2DM is doubled in patients with NAFLD [87]. Moreover, 25% of patients with NAFLD also have T2DM [88].
NAFLD and insulin resistance are interconnected, and therefore the development of prediabetes and diabetes is the most direct consequence of them at the extrahepatic level [11,89]. Particularly, the coexistence of NAFLD and T2DM worsens the course of both conditions since this relationship is bidirectional [90][91][92]. One meta-analysis found that the risk of T2DM is greater in patients with advanced NAFLD with fibrosis [84], and in another meta-analysis evaluating whether NAFLD predicted T2DM, NAFLD predicted the risk of T2DM independent of age and obesity, irrespective of the NAFLD diagnosis method (ultrasonography or elevated liver enzymes) [85]. Moreover, this observation was also applicable to patients with prediabetes and NAFLD, where the incidence of T2DM was higher than in patients with prediabetes without NAFLD [86]. When different types of diabetes were analyzed, the prevalence of NAFLD was low in patients with type 1 diabetes mellitus but high in T2DM patients in whom NAFLD was associated with increasing BMI, triglycerides, ALT (alanine aminotransferase) serum concentration, and decreasing adiponectin concentration [93]. Surprisingly, NAFLD occurred more often in T2DM patients not treated with insulin than in patients treated with insulin [93].
In another cohort study of 10,141 participants, future diabetes mellitus risk could be modified with time by changes in NAFLD status where resolution of NAFLD could diminish the risk of diabetes onset. On the other hand, the development of NAFLD raised the risk of developing diabetes [94]. Current evidence suggests that the magnitude of risk of incident T2DM mirrors the severity of NAFLD, especially with the severity of liver fibrosis [95].
It is important to distinguish between simple steatosis, which does not progress to advanced fibrosis, and non-alcoholic steatohepatitis (NASH), which can lead to hepatocellular carcinoma. Indeed, NASH is characterized by fat accumulation, inflammation and necrosis, ballooning of the cells, and different stages of liver fibrosis, up to cirrhosis. The only method to differentiate between steatosis and NASH is liver biopsy. Masarone et al. performed this invasive procedure in 215 patients with elevated transaminases and metabolic syndrome or T2DM. The prevalence of NAFLD in patients with metabolic syndrome was 94.82% and was present in all the patients with T2DM. NASH was found in 58.52% of participants with metabolic syndrome and 96.82% of T2DM patients. According to the authors, one can assume that patients with T2DM have NASH. As insulin resistance is of crucial importance in the pathophysiology of both T2DM and NASH, NASH may be one of the early complications of T2DM [96].
Some studies have reported that a reduction in the T2DM incidence [97] and the clinical manifestation of atherosclerosis [98] could be achieved by the pharmacological eradication of the hepatitis C virus with direct-acting antiviral drugs [99], caused by the reduction of insulin resistance and improving various HCV-induced glucose homeostasis mechanisms [97,98].
The strong link between T2DM and NAFLD was underlined in 2020 by an international panel of experts who proposed the new term MAFLD instead of NAFLD [10]. The results of the studies described above are presented in Table 4. Abbreviations: CI-credible interval, DM-diabetes mellitus, HR-hazard ratio, NAFLD-non-alcoholic fatty liver disease, NASH-non-alcoholic steatohepatitis, RR-risk ratio, T1DM-type 1 diabetes mellitus, T2DM-type 2 diabetes mellitus.

Treatment of NAFLD with Antidiabetic Drugs
Currently, there is no single, independent of the presence or absence of T2DM, pharmaceutical treatment for NAFLD that has been approved by international guidelines [9]. The frequent coexistence of diabetes and NAFLD and complex pathogenesis of these two metabolic diseases result in the growing interest in antidiabetic drugs used in the treatment of NAFLD.
Current guidelines state that pharmacotherapy is reserved for patients with NASH and for patients at high risk of disease progression. Among the antidiabetic drugs recommended, only pioglitazone is used for the treatment of NASH with insulin resistance (evidence level A, strength 2) [9,124]. Older studies showed that NASH treatment with pioglitazone caused histological improvement of hepatic steatosis and fibrosis [100][101][102][103].
In an RCT analyzing patients with T2DM or prediabetes and NASH, pioglitazone usage for 18-months resulted in a reduction of fibrosis score and hepatic triglyceride content and improved insulin sensitivity of adipose tissue, liver, and muscles [103]. In a recent meta-analysis, pioglitazone treatment in patients with NAFLD with or without T2DM caused significant reductions of ALT and AST (aspartate aminotransferase) levels [104].
An upcoming drug of huge interest is a dual GIP and GLP-1 RA agent (tirzepatide), which is under investigation. It is being administered once a week in the therapy of T2DM patients with NASH and fibrosis [123]. Treatment with this drug for 26 weeks compared to dulaglutide and placebo resulted in a significant reduction in NASH-related biomarkers, an increase in adiponectin, and a greater reduction of ALT, compared to dulaglutide treatment [123]. A published network meta-analysis of RCTs [126] assessed the effectiveness of antidiabetic medications for T2DM as potential therapeutic agents for NAFLD, comparing SGLT-2i, GLP-1 RA, PPARγ (peroxisome proliferator-activated receptor) agonists, biguanides, sulfonylureas, and insulin. This showed that GLP-1 RA and SGLT-2i led to reductions in BMI, fibrosis, and steatosis, with SGLT-2i being the best treatment for reducing low-density lipoprotein cholesterol (LDL-C) and rising HDL-C. Studies related to the SGLT-2i, GLP-1 RA, and dual GIP and GLP-1-RA use in the treatment of NAFLD are summarized in Table 5.  Pioglitazone had a significant effect on the levels of ALT and AST but was also associated with an increased risk of weight gain and increased BMI. Liraglutide and metformin had significant effects on reducing ALT and AST. Abbreviations: ALT-alanine aminotransferase, AST-aspartate aminotransferase, BMI-body mass index, CAPcontrolled attenuation parameter, CI-credible interval, IHF-intrahepatic fat, GLP-1 RA-glucagon-like peptide 1 receptor agonists, GGT-gamma-glutamyl transferase, HbA 1c -glycated haemoglobin A1c, HDL-c-high density lipoprotein cholesterol, HOMA-IR-homeostatic model assessment for insulin resistance, LDL-c-low density lipoprotein cholesterol, LFC-liver fat content, LSM-liver stiffness measurement, MD-mean difference, mgmilligram, MRI-magnetic resonance imaging, MRI-PDFF-magnetic resonance imaging derived proton density fat fraction, NAFLD-non-alcoholic fatty liver disease, NASH-non-alcoholic steatohepatitis, OR-odds ratio, PPARγ-peroxisome proliferator-activated receptor, RCT-randomized controlled trial, RR-relative risk, SATsubcutaneous adipose tissue, SGLT-2i-sodium-glucose cotransporter type-2 inhibitors, SMD-standardized mean difference, T2DM-type 2 diabetes mellitus, TAG-triglycerides, TC-total cholesterol, VAT-visceral adipose tissue, WMD-weighted mean difference.

NAFLD and Cardiovascular Disease
Both NAFLD and CVD are highly prevalent and associated with metabolic disturbances; hence they frequently coexist [127]. This causal relationship may be due to shared common pathophysiological pathways, which include low-grade inflammation, oxidative stress, and insulin resistance [128].
NAFLD is linked with different manifestations of CVD, including subclinical atherosclerosis, overt atherosclerosis, and cardiovascular events and deaths [24,[129][130][131]. Hence, both the American Association for the Study of Liver Disease and the European Association for the Study of the Liver suggest cardiovascular screening in patients with NAFLD [9,132]. In relation to the association of NAFLD with the risk of major adverse cardiovascular events (MACE), several meta-analyses and cohort studies show that in patients with NAFLD, the risk of MACE is increased, independent of other cardiovascular risk factors or the extent of coronary disease [133,134].
One meta-analysis from the year 2021 deserves special attention because it was performed among people with histologically confirmed NAFLD who did not present with CVD at baseline (10,422 participants) and were prospectively followed-up for a median of 13.6 years [134]. This study showed that patients with biopsy-proven NAFLD who were matched to controls had a higher incidence of MACE, which included ischemic heart disease, congestive heart failure, and cardiovascular mortality [134]. Moreover, the rates of incident MACE increased progressively with worsening NAFLD severity.
There are also studies that indicated that CVD risks in patients with NAFLD who are non-obese [135] or non-overweight [136] were also increased. Additionally, in obese patients with NAFLD, liver fat content (LFC) >10% was also a predictor of subclinical atherosclerosis [135].
One retrospective, "real world" cohort study aimed to describe the CVD burden and mortality in patients with NAFLD during 14-years follow-up observation following hospital discharge, showing that in patients with non-cirrhotic NAFLD, the condition was associated with increased overall mortality [137]. In contrast, other studies have stated that NAFLD was not correlated with CVD mortality [138][139][140] nor an increased risk of acute myocardial infarction or stroke [141].
Due to the recent change in nomenclature of fatty liver disease (NAFLD to MAFLD), the prevalence of fatty liver disease and associated CVD risk required re-evaluation, taking into account each of these definitions separately [142]. An analysis of 9,584,399 participants aged 40-64 from the National Health Database revealed that MAFLD and NAFLD, independent of the definition used, increased the risk of CVD events [142]. Furthermore, CVD seems to be increased when NAFLD coexists with T2DM. A meta-analysis of 11 studies, including cross-sectional and cohort studies, indicated that the risk for CVD in T1DM and T2DM patients with NAFLD was increased two-fold compared to patients without NAFLD [143]. In T2DM patients with NAFLD, the occurrence of coronary artery disease (CAD) was more prevalent than in T2DM patients without NAFLD [144][145][146]. Interestingly, patients diagnosed with NAFLD before T2DM diagnosis showed a higher prevalence of CAD and hypertension when compared to groups of patients with T2DM without NAFLD and T2DM diagnosed before NAFLD [144]. Additionally, T2DM patients with NAFLD had a higher prevalence of hypertension, obesity, higher hemoglobin A1c (HbA1c) levels, higher triglyceride, lower HDL-C concentration, and higher mean carotid IMT (intima-media thickness) [145].
On the other hand, another study concerning atherosclerotic lesions in T2DM patients found no significant difference in carotid IMT between T2DM patients with and without NAFLD [147]. The same study demonstrated that the prevalence of carotid and lower limb atherosclerotic plaque and stenosis was higher in T2DM patients with NAFLD compared to T2DM patients without NAFLD [147]. A study by Kim et al. divided T2DM patients into two groups: insulin-resistant and insulin-sensitive, whereby mean carotid IMT was higher in subjects with both insulin resistance and NAFLD than in insulin-sensitive patients with or without NAFLD [148]. T2DM patients with co-existing NAFLD also have an increased risk of PAD (peripheral artery disease) defined in this study as ABI (ankle-brachial index) < 0.90 on either side [149].
While an association between NAFLD and T2DM microvascular complications seems plausible, the number of studies on this topic is limited and inconclusive [144,[150][151][152][153][154]. In 2008, Targher et al. performed one of the first studies evaluating the associations between NAFLD and both chronic kidney disease (CKD) and retinopathy in a large cohort of 2103 patients with T2DM and found that NAFLD was independently associated with an increased prevalence of CKD and retinopathy [151]. In the Valpolicella Heart Diabetes Study, among 1760 outpatients with T2DM who had normal kidney function at baseline, those with NAFLD had an independently increased risk of incident CKD over a follow-up period of 6.5 years [155]. These initial observations have been confirmed in further clinical studies and meta-analysis of observational studies [152,153], and moreover, it was shown that NAFLD increases the risk of diabetic neuropathy [156]. However, there are also studies that do not show such a relationship [144,150,154]. Results of the relevant studies are included in Table 6.
It is well established that patients with DM compared to people without carbohydrate disorders have a higher risk of heart failure and CVD. There is a term "diabetic cardiomyopathy", deteriorating patient's prognosis and being described as a form of heart disease occurring in diabetic patients, which causes significant structural as well as functional changes in the myocardium. There is a common pathophysiological mechanism of diabetic cardiomyopathy and NAFLD, namely insulin resistance [157]. It results in an increase in lipogenesis in the liver and adipose tissue's lipolysis inhibition [89]. Insulin resistance also causes a decrease in the concentration of an insulin-sensitizing adipokine called adiponectin. Physiologically, adiponectin plays an important role in hepatoprotection by modifying free fatty acids metabolism, gluconeogenesis, and lipogenesis. Moreover, by reducing the number of proinflammatory cytokines and promoting the proliferation of hepatic stellate cells, it prevents liver fibrosis. In NAFLD, there are a couple of mechanisms (including dysfunction in adipokine production, increase in oxidative stress reactions, and general proinflammatory state) that influence the atherosclerotic plaque formation and its progression, therefore, leading to the increase of cardiovascular risk [158]. NAFLD was associated with an increased risk of prevalent and incident CKD. NASH was associated with a higher prevalence and incidence of CKD than simple steatosis. Advanced fibrosis was associated with a higher prevalence and incidence of CKD than non-advanced fibrosis.

Mellinger et al. (2015) [131]
Prospective cohort 1/3014 3 years Adults with or without NAFLD There was no significant association between NAFLD and CVD. NAFLD was associated with CAC and AAC.

Conclusions
MAFLD is a new clinical definition for fatty liver disease, which shifts NAFLD from a disease of exclusion to one of inclusion, where the pathogenic processes originate from underlying metabolic dysfunction. Because MAFLD is not widely used terminology in the scientific literature, most published data focus on NAFLD. The latter as an epidemic is tightly linked to T2DM, which are known to frequently coexist with and synergistically increase the CVD risk.
Despite the high prevalence of NAFLD and many epidemiological studies showing correlations between NAFLD and CVD, it is still difficult to unequivocally identify a causal relationship between the two entities [162] and to show that NAFLD is an independent risk factor for CVD [163], given the presence of many comorbidities and confounding factors. In addition, the available studies show great heterogeneity. Also, the genetic variants that predispose to the development of NAFLD have not been linked to the development of atherosclerotic CVD in the absence of general obesity and metabolic syndrome [164].
We are still unclear whether the diagnosis of NAFLD can be used as a tool to improve cardiovascular risk and modify treatment [162]. Lifestyle interventions are recommended by the European clinical guidelines as the best therapeutic option for human NAFLD [9,165]. Moreover, ≥7% weight loss improves steatosis significantly, resulting in lowering of the NAFLD activity score (NAS) [166,167]. On the other hand, only 40% of patients in the above study reached that goal and reduced steatohepatitis, underlining the difficulties in managing NAFLD with lifestyle changes [166]. Nevertheless, reduction of fructose should be recommended for patients with NAFLD along with emerging therapies that can lower the activity of liver enzymes, fibrosis, and inflammation, such as PPARγ inhibitors (pioglitazone), SGLT-2i, and GLP-1 RA, as well as modification in gut microbiota.

Data Availability Statement:
No new data were created or analyzed in this study. Data sharing is not applicable to this article.

Conflicts of Interest:
The authors declare no conflict of interest directly related to this paper.